This invention relates to a process for preparing a radiopharmaceutical diagnostic agent useful in scanning applications of organs and tissues in a mammal. More particularly, the invention relates to a sterile, stable macroaggregates of human serum albumin suspended in a highly purified, stable human serum albumin carrier in a form which is labeled with technetium-99mTc prior to injecting the human serum albumin into a patient for diagnostic scanning of the lung.
Macroaggregated particles of human serum albumin having a particle size of 3 to 150 microns in diameter radiolabeled with technetium-99m have been used in lung function studies and diagnosis. After injection into the patient, the radiolabeled macroaggregated particles diffuse into and are retained by the capillary system of the lung allowing scintigraphic visualization of the vascular system by a diagnostic practitioner. The radiolabeled macroaggregates obstruct the capillaries for only a short time and cause no harm to the pulmonary system. Subsequent to the short retention, the macroaggregates are digested by the phagocytes in the blood thereby clearing the capillaries from the macroaggregates. Technetium-99m, which emits gamma rays, has a radioactive half-life of only about 6 hours and the associated radiation disappears along with the particles of the macroaggregates from the pulmonary system.
A critical characteristic in macroaggregated human serum-containing product is particle size. The product is designed for the macroaggregates to be trapped in the pulmonary capillaries. If embolisms are present in the capillary system, the macroaggregates will not travel through the blockage indicating blood circulatory problems by the lack of radiation from the area of the capillaries where the macroaggregate particles could not enter.
Such blockage may also be caused by macroaggregates too large to pass through certain portions of healthy capillaries thereby indicating inadequate blood circulation or disease. Macroaggregated particles larger than 150 microns in diameter may even block large capillaries. On the other hand, when the macroaggregated particles are smaller than about 3 microns in diameter, they pass through the walls of the capillaries directly into the liver causing shadows and distort the scintiphoto of the lung. (See, for example, U.S. Pat. No. 3,987,157.) To eliminate the problem of artificial blockage of capillaries by large particle size macroaggregates and to prevent passing of small particle size aggregates, the prior art sets a limit on particle size range of about 5 to 10 micron minimum and of about 80 to 100 micron maximum average diameter, while the preferred range is about 15 to 30 micron minimum and 30 to 50 micron maximum in diameter. (See, for example, U.S. Pat. Nos.: 3,674,900; 3,863,004; 3,862,299; 4,024,233; and 4,094,965.)
To obtain macroaggregates of the required particle size, the prior art uses a heating step the temperature of which ranges of from about 70xc2x0 to 100xc2x0 C., and a cooling step ranging from about 18xc2x0 to 22xc2x0 C.
Although the particle size of the macroaggregates will be in the desired range, the distribution of particle size will vary resulting in a mixture of small, medium and large particles. When radiolabeled, the particles will not carry an even amount of technetium-99m resulting in reduced diagnostic efficacy. Also, the blood flow in the capillaries will preferentially carry the smaller particles, leaving behind the larger particles which travel at a slower rate of speed. This results in a non-uniform distribution of the technetium-99m which influences the scintigraphic reading of the condition of capillaries and tissues. It is, therefore, important to produce macroaggregated particles of essentially uniform size so that the radiolabeled particles will be evenly distributed in the capillaries and tissues and give a reliable reading of maximum diagnostic efficacy on the condition of the pulmonary system.
The prior art also encountered the problem of agglomeration of the particles. U.S. Pat. No. 3,863,004 discloses the labeling of denatured albumin with technetium sulfur colloid which tended to agglomerate on standing and required ultrasonic treatment of the particles prior to use. U.S. Pat. No. 4,187,285 uses an anti-agglomerating surfactant, such as Poloxamer 188, to guard against agglomeration. The ultrasonic treatment suggested in the former patent is cumbersome, while the use of a surfactant suggested in the latter patent is less desirable than a radiolabeled human serum albumin without additives such as a surface active agent.
Commercially supplied albumin is a 25% albumin solution in an aqueous diluent containing additives such as sodium carbonate, sodium citrate and/or acetyltryptophan.
We have discovered that to obtain an efficacious albumin product the commercially available human albumin solution must first be purified prior to proceeding with the aggregating step.
We have also discovered that a technetium-99m labeled human serum albumin product can be made essentially without residual amounts of sodium acetate/acetic acid buffer and hydrochloric acid which are used in the prior art processes.
We have further discovered that particle size can be made essentially uniform by using a controlled heating-cooling cycle when producing the macroaggregates. This enables the control of the number of particles per vial.
We have also discovered that the desired particle size can be further assured by passing the macroaggregates through a size restricting screen to meet USP requirements.
The process for preparing the injectable suspension of the present invention comprises the steps of:
a) mixing human serum albumin with 0.9% w/v saline solution and ultrafiltering the mixture through a membrane having a porosity of 10,000 nominal molecular weights limit;
b) mixing a buffer having a pH of from 4.95 to 5.25 and adding the buffer to the ultrafiltered mixture to obtain a buffered, ultrafiltered mixture;
c) filtering through a sterile 0.2 micron filter about a 0.43% w/v stannous chloride in HCl solution and adding the solution to the buffered, ultrafiltered mixture to obtain a reaction mixture;
d) heating the reaction mixture gradually so that it attains about 75xc2x0 C. to 95xc2x0 C. in about 90 to 100 minutes;
e) cooling the reaction mixture gradually thereby entrapping particles of stannous chloride in the macroaggregates so formed;
f) separating the macroaggregates and resuspending them in sterile water for injection;
g) filtering the resuspended macroaggregates through a 75 micron sterile sizing screen;
h) determining the concentration of the human serum albumin;
i) adjusting the concentration of human serum albumin to the desired dose;
j) transferring the desired volumes into vials and lyophilizing their content; and
k) reconstituting the lyophilized human serum albumin with an aqueous solution of 99mTc pertechnate or another isotope of 99mTc such as Tc94 or Tc96.
The injectable suspension of the present invention is intravenously administered to a patient for determining the pulmonary blood supply. Following the intravenous administration the 99mTc macroaggregated albumin are trapped in functioning pulmonary capillaries and small arterioles. An image of in-vivo radioactivity distribution will then shown the pulmonary lung perfusion.
Although the description herein concerns the use of human serum albumin to form the macroaggregates of this invention, other proteins may be utilized such as, for example, alpha, beta or gamma globulin or fibrinogen.